Francisella tularensis, the etiological agent of tularemia, is one of the most infectious bacterial pathogens currently known. Tularemia can be a fatal disease in humans and although outbreaks are thought to be rare, infections caused by F. tularensis have become a public health concern because of the potential for use of this organism as a biological weapon. Evidence suggests that the pathogenicity of F. tularensis depends on its ability to survive and replicate within macrophages; nevertheless, little is known about the detailed molecular mechanisms underlying F. tularensis pathogenesis. This proposal focuses on MgIA, a putative regulator of virulence gene expression that is essential for F. tularensis to survive within macrophages. Our ultimate goal is to understand the molecular mechanism by which MgIA controls virulence gene expression in F. tularensis. [unreadable] The first aim of this proposal is to identify those genes that constitute the MgIA regulon in F. tularensis using DNA microarrays; virulence genes that are required for intramacrophage growth and survival are expected to be amongst this set. The second and third aims are to investigate the mechanism by which MgIA regulates the expression of target genes. In aim two we will use a combination of biochemical and genetic approaches to determine whether a second MglA-like protein we have uncovered in F. tularensis also influences the expression of MglA-regulated genes. In aim three we will test a specific hypothesis concerning MgIA function by seeking to identify proteins that bind to putative MglA-dependent promoters and are required for the MglA-dependent expression of target genes. As well as facilitating the design of novel antimicrobial agents aimed at inhibiting MgIA function, our studies will provide several genetic and biochemical tools for furthering the understanding of F. tularensis pathogenicity. [unreadable] [unreadable] [unreadable]